Preparation and Evaluation of Lipid Emulsion Containing 13 Vitamins for Injection Without Anaphylactoid Reactions.

OBJECTIVE: Multivitamins containing Tween 80 can cause anaphylactoid reactions. The objective of this study was to develop a new lipid emulsion containing 13 fat- and water-soluble vitamins for injection (13V-LE) that were simultaneously dissolved in one bottle and to evaluate the stability of and anaphylactoid reactions to 13V-LE. METHODS: Particle size, ζ-potential, and polydispersity of 13V-LE were assayed with a Zetasizer Nano ZS. Entrapment efficiency of 13V-LE was determined with HPLC. Behavior, histamine, and blood pressure of beagle dogs were investigated by observation, fluorospectrophotometry, and sphygmomanometry. RESULTS: The 13V-LE with the smallest particles and highest entrapment efficiency with stable ζ-potential was composed of soybean oil, glycerin (2.25%, w:v), egg lecithin (1.2%, w:v), and purified water. There was no obvious change in characteristics of the 13V-LE samples in terms of appearance, size distribution, ζ-potential, pH value, or concentration over 6 months. In anaphylactoid reactions tests, when being administered with the multivitamin Infuvite Adult containing Tween 80, six beagles showed grade IV symptoms (P<0.01 vs control), low blood pressure, and high plasma-histamine concentrations (P<0.05 or P<0.01). However, there were no significant differences in behavior, blood pressure, or histamine concentration in the dogs before and after administration in the 13V-LE group. CONCLUSION: The 13V-LE formulation is a suitable intravenous lipid emulsion without anaphylactoid reactions.

Ligands and Signaling of Mas-Related G Protein-Coupled Receptor-X2 in Mast Cell Activation.

Mas-related G protein-coupled receptor-X2 (MRGPRX2) is known as a novel receptor to activate mast cells (MCs). MRGPRX2 plays a dual role in promoting MC-dependent host defense and immunomodulation and contributing to the pathogenesis of pseudo-allergic drug reactions, pain, itching, and inflammatory diseases. In this article, we discuss the possible signaling pathways of MCs activation mediated by MRGPRX2 and summarize and classify agonists and inhibitors of MRGPRX2 in MCs activation. MRGPRX2 is a low-affinity and low-selectivity receptor, which allows it to interact with a diverse group of ligands. Diverse MRGPRX2 ligands utilize conserved residues in its transmembrane (TM) domains and carboxyl-terminus Ser/Thr residues to undergo ligand binding and G protein coupling. The coupling likely initiates phosphorylation cascades, induces Ca2+ mobilization, and causes degranulation and generation of cytokines and chemokines via MAPK and NF-κB pathways, resulting in MCs activation. Agonists of MRGPRX2 on MCs are divided into peptides (including antimicrobial peptides, neuropeptides, MC degranulating peptides, peptide hormones) and nonpeptides (including FDA-approved drugs). Inhibitors of MRGPRX2 include non-selective GPCR inhibitors, herbal extracts, small-molecule MRGPRX2 antagonists, and DNA aptamer drugs. Screening and classifying MRGPRX2 ligands and summarizing their signaling pathways would improve our understanding of MRGPRX2-mediated physiological and pathological effects on MCs.

Estimating the instant case fatality rate of COVID-19 in China.

OBJECTIVE: The outbreak of coronavirus disease 2019 (COVID-19) in China has been basically controlled. However, the global epidemic of COVID-19 is worsening. We established a method to estimate the instant case fatality rate (CFR) and cure rate of COVID-19 in China. METHODS: A total of 82 735 confirmed cases released officially by the Chinese authorities from December 8, 2019 to April 18, 2020 were collected. The estimated diagnosis dates of deaths and cured cases were calculated based on the median cure time or median death time of individual cases. Following this, the instant CFR was calculated according to the number of deaths and cured cases on the same estimated diagnosis date. RESULTS: In China, the instant CFR of COVID-19 was 3.8-14.6% from January 1 to January 17; it then declined gradually and stabilized at 5.7% in April. The average CFR in China was 6.1±2.9%, while the CFR was 1.0±0.4% in China except Hubei Province. The cure rate of COVID-19 was 93.9±2.9% in China, and stabilized at 94.3%, while it was 99.0±0.4% in China except Hubei Province. CONCLUSIONS: The instant CFR of COVID-19 in China overall was much higher than that in China except Hubei Province. The CFR of COVID-19 in China was underestimated.

Determination of pharmacokinetic parameters of vitamin K1 in rats after an intravenous infusion.

Pharmacokinetic parameters of vitamin K1 have a large range of values in different literature. The aim of this study was to determine the pharmacokinetic parameters of vitamin K1 following post-constant speed intravenous infusion (PCSII) to provide rational pharmacokinetic parameters of vitamin K1 and compare these with results of noncompartmental analysis following intravenous injection (IV). After 15 hours intravenous infusion of vitamin K1 in rats, the logarithmic concentration-time curve of vitamin K1 was fit to a linear equation following PCSII (R2  = 0.9599 ± 0.0096). Then, half-time (T1/2 ), apparent volume of distribution (Vd ), and clearance rate (CL) were estimated successively. T1/2 of vitamin K1 was 4.07 ± 0.41 hour, CL was 89.47 ± 3.60 mL/h, and Vd was 525.38 ± 54.45 mL in rats following PCSII. There was no significant difference in pharmacokinetic parameters of vitamin K1 among different sampling times. For noncompartmental analysis, T1/2 and mean residence time (MRTINF ) for a sampling duration of 6h were shorter than those of 12 hours or 24 hours sampling duration following IV (P < .05, P < .01). In addition, T1/2 of vitamin K1 was obviously different from MRT-equated half-time (T1/2,MRT )(P < .05). Vd and CL of vitamin K1 following PCSII were larger than those following IV based on noncompartmental analysis (P < .01). The results demonstrated that drug distribution in the body was balanced and the Napierian logarithmic concentration-time curve of vitamin K1 fit to a linear equation following PCSII. Vitamin K1 has a long T1/2 and a relatively large Vd following PCSII.

Hydrogen Peroxide-Mediated Oxygen Enrichment Eradicates Helicobacter pylori In Vitro and In Vivo.

Helicobacter pylori is an important risk factor for gastric ulcers. However, antibacterial therapies increase the resistance rate and decrease the eradication rate of H. pylori Inspired by the microaerophilic characteristics of H. pylori, we aimed at effectively establishing an oxygen-enriched environment to eradicate and prevent the recurrence of H. pylori The effect and the mechanism of an oxygen-enriched environment in eradicating H. pylori and preventing the recurrence were explored in vitro and in vivo During oral administration and after drug withdrawal, H. pylori counts were evaluated by Giemsa staining in animal cohorts. An oxygen-enriched environment in which H. pylori could not survive was successfully established by adding hydrogen peroxide into several solutions and rabbit gastric juice. Hydrogen peroxide effectively killed H. pylori in Columbia blood agar and special peptone broth. Minimum inhibition concentrations and minimum bactericidal concentrations of hydrogen peroxide were both relatively stable after promotion of resistance for 30 generations, indicating that hydrogen peroxide did not easily promote resistance in H. pylori In models of Mongolian gerbils and Kunming mice, hydrogen peroxide has been shown to significantly eradicate and effectively prevent the recurrence of H. pylori without toxicity and damage to the gastric mucosa. The mechanism of hydrogen peroxide causing H. pylori death was related to the disruption of bacterial cell membranes. The oxygen-enriched environment achieved by hydrogen peroxide eradicates and prevents the recurrence of H. pylori by damaging bacterial cell membranes. Hydrogen peroxide thus provides an attractive candidate for anti-H. pylori treatment.

Label-Free Quantitative Proteomic Profiling of LAD2 Mast Cell Releasates Reveals the Mechanism of Tween-80-Induced Anaphylactoid Reaction.

PURPOSE: Tween-80 is one of the most important causes resulting in anaphylactoid reaction. However, its mechanism remains unclear. Proteomic characterizations of mast cells' excreta in response to Tween-80 are assayed to investigate the mechanism of anaphylactoid reaction. EXPERIMENTAL DESIGN: A label-free LCMS/MS-based proteomics is used to analyze Tween-80-stimulated Laboratory of Allergic Diseases 2 (LAD2) mast cells releasates. The results of proteomic are analyzed by bioinformatics analysis. Western blotting is used to verify the expression of proteins. RESULTS: Overall, endocytosis, nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB), and calcium signaling pathways play important roles in Tween-80-induced LAD2 cells activation by bioinformatics analysis. The expressions of relative proteins including actin-related protein 2/3 complexes, vacuolar protein sorting-associated protein, phosphorylation of transcription factor of P105 and P65, phosphorylation of inositol 1,4,5-trisphosphate receptor (IP3 R), phosphoinositide phospholipase Cγ (PLCγ), and protein kinase C (PKC), are significantly increased in Tween-80 group compared to control. Tween-80 might be internalized via endocytosis, which induces degranulation by PLCγ/PKC pathways mediated calcium influx, and promotes the generation of inflammatory mediators via NF-κB pathway resulting in anaphylactoid reaction.

Non-IgE-mediated hypersensitivity induced by multivitamins containing Tween-80.

Multivitamins have been widely used for years. Adverse reactions, especially hypersensitivity, to multivitamins are becoming noteworthy. However, the classification of hypersensitivity is confusing, and the trigger is unknown. Multivitamins consist of two vials labelled vial 1 containing Tween-80 and vial 2. Multivitamins without Tween-80 were used as a contrast. Behaviouristics, histamine, IgE, and blood pressure of beagle dogs and guinea-pigs were investigated by observation, ELISA and sphygmomanometer, and degranulation and apoptotic of RBL-2H3 cells were assayed by spectrophotometry and flow cytometry. The results showed that dogs suffered from multiorgan anaphylactoid symptoms, and dramatically decreased blood pressure, and high plasma concentrations of histamine after the first administration of multivitamins and multivitamins vial 1, which contains Tween-80, compared to the control, multivitamins vial 2 or multivitamins without Tween-80. In anaphylaxis assay, guinea-pigs did not display any anaphylaxis symptoms and there were no changes in plasma histamine and IgE concentrations in the multivitamins and multivitamins vial 1 groups or in the multivitamins vial 2 and multivitamins without Tween-80 groups except ovalbumin. Compared to the control, the release of ß-hexosaminidase and histamine, and the apoptosis of non-antigen-sensitized RBL-2H3 cells significantly increased in the Tween-80 and multivitamins and multivitamins vial 1 groups in a concentration-dependent manner. However, there was no alteration in multivitamins vial 2 and multivitamins without Tween-80 groups. The results indicate that the hypersensitivity induced by multivitamins may be anaphylactoid reaction, but not anaphylaxis. Multivitamin-induced release of inflammatory factors is triggered by Tween-80 through a non-IgE-mediated pathway.

Crocin alleviates coronary atherosclerosis via inhibiting lipid synthesis and inducing M2 macrophage polarization.

Atherosclerosis is a chronic inflammatory disease arising from an imbalance in lipid levels and the accumulation of cholesterol-laden macrophages in the artery wall. Crocin is an active ingredient of Crocus sativus L. This study established a rat coronary atherosclerosis model induced by vitamin D3 (VD3), to explore the effect of Crocin on lipid metabolism, macrophage polarization and the activity of inflammatory proteins. The results revealed that Crocin decreased blood lipid levels by decreasing the levels of endothelin (ET), total cholesterol (TC), triglyceridelow (TG) and low-density lipoprotein cholesterol (LDL-c), elevating the level of high-density lipoprotein cholesterin (HDL-c). Crocin also inhibited lipogenesis by suppressing the expression of lipogenesis-related proteins and elevating lipid catabolism-related proteins. Moreover, Crocin effectively alleviated inflammation by suppressing the expression of pro-inflammatory cytokines and increasing levels of anti-inflammatory cytokines. We further found that Crocin promoted macrophage polarization to the M2 phenotype by reducing M1 markers (CD40+ and CD11c+) and elevating M2 markers (CD68+ and CD206+). Finally, Crocin strongly inhibited the expression of NF-κB p65 and its translocation into the nucleus. Crocin partially counteracted nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) p65 expression and the nuclei accumulation caused by NF-κB p65 overexpression. Taken together, our research indicated that Crocin inhibited lipogenesis and alleviated the inflammation in a VD3-induced rat coronary atherosclerosis model by promoting M2 macrophage polarization and maybe by inhibiting NF-κB p65 nuclear translocation. This study implicates Crocin as a potential therapeutic strategy for coronary atherosclerosis.

Finding the optimal dose of vitamin K1 to treat vitamin K deficiency and to avoid anaphylactoid reactions.

Vitamin K1 injection induces severe dose-related anaphylactoid reactions and overdose for the treatment of vitamin K deficiency. We aimed to find an optimal and small dose of vitamin K1 injection to treat vitamin K deficiency and avoid anaphylactoid reactions in animal. Rats were administered a vitamin K-deficient diet and gentamicin to establish vitamin K deficiency model. Behaviour tests were performed in beagle dogs to observe anaphylactoid reactions. The results showed an increased protein induced by vitamin K absence or antagonist II (PIVKA-II) levels, a prolonging of prothrombin time (PT) and activated partial thromboplastin time (APTT) and a decrease in vitamin K-dependent coagulation factor (F) II, VII, IX and X activities in the model group. In vitamin K1 0.01 mg/kg group, the liver vitamin K1 levels increased fivefold and the liver vitamin K2 levels increased to the normal amount. Coagulation markers PT, APTT, FVII and FIX activities returned to normal. Both in the 0.1 and 1.0 mg/kg vitamin K1 groups, coagulation functions completely returned to normal. Moreover, the amount of liver vitamin K1 was 40 (0.1 mg/kg) or 100 (1.0 mg/kg) times as in normal. Vitamin K2 was about 4 (0.1 mg/kg) or 5 (1.0 mg/kg) times as the normal amount. There was no obvious anaphylactoid symptom in dogs with the dose of 0.03 mg/kg, which is equivalent to the dose of 0.01 mg/kg in rats. These results demonstrated that a small dose of vitamin K1 is effective to improve vitamin K deficiency and to prevent anaphylactoid reactions, simultaneously.

H2S Prevents Cyclosporine A-Induced Vasomotor Alteration in Rats.

Cyclosporine A (CsA) induces hypertension after transplantation. Hydrogen sulfide (H2S) was found to have hypotensive/vasoprotective effects in the cardiovascular system. The present study aims to investigate the role of H2S on CsA-induced vascular function disorder in rats. Rats were subcutaneously injected with CsA 25 mg/kg for 21 days. Blood pressure was measured by the tail-cuff method. Vasomotion was determined using a sensitive myograph. Western blotting and immunohistochemistry were used to quantify the protein expression of endothelin type A (ETA) receptor and essential MAPK pathway molecules. Vascular superoxide anion production and serum contents of malondialdehyde were determined. The results showed that sodium hydrosulfide (NaHS), a H2S donor, significantly attenuated the increase of blood pressure and contractile responses, and the upregulation of ETA receptor induced by CsA. In addition, NaHS could restore the CsA decreased acetylcholine-induced vasodilatation. Furthermore, NaHS blocked the CsA-induced elevation of reactive oxygen species level, extracellular signal-regulated kinase and p38 MAPK activities. In conclusion, H2S prevents CsA-induced vasomotor dysfunction. H2S attenuates CsA-induced ETA receptor upregulation, which may be associated with MAPK signal pathways. H2S ameliorates endothelial-dependent relaxation, which may be through antioxidant activity.

The expression of bitter taste receptors in mesenteric, cerebral and omental arteries.

AIM: Bitter taste is sensed by the bitter taste receptor (TAS2R), which is mainly expressed in the tongue as well as in extra-oral organs, such as the gastrointestinal tract, respiratory tract, brain, heart and testis. This study aimed to investigate whether TAS2R is expressed in the mesenteric, cerebral and omental arteries. MAIN METHODS: The expression levels of TAS2R mRNA and protein were determined by reverse-transcription polymerase chain reaction and Western blotting, respectively. The location of TAS2R was determined by immunofluorescence imaging. TAS2R agonists were used in a sensitive myograph to study the function of TAS2R in arteries. KEY FINDINGS: The mRNA of rat TAS2Rs, including rTAS2R39, rTAS2R40, rTAS2R108, rTAS2R114, rTAS2R130, rTAS2R137, and rTAS2R140, was expressed in rat mesenteric and cerebral arteries, but rTAS2R114 was not expressed in the cerebral arteries. The mRNA of human TAS2Rs, including hTAS2R3, hTAS2R4, hTAS2R7, hTAS2R10, hTAS2R14, hTAS2R39 and hTAS2R40, was expressed in human omental arteries. The TAS2R7 protein was expressed in rat mesenteric and cerebral arteries, as well as in human omental arteries. Immunofluorescence imaging confirmed that TAS2R7 was located in vascular smooth muscle cells and endothelial cells. The TAS2R agonists, chloroquine and quinine relaxed rat mesenteric arteries and cerebral arteries and human omental arteries in a concentration-dependent manner. SIGNIFICANCE: TAS2R is expressed in the arteries of systemic circulation, including rat mesenteric and cerebral arteries and human omental arteries. This study provides evidence that TAS2R do exist in the arteries and may be involved in the mediation of vessel functions.

H2S induces vasoconstriction of rat cerebral arteries via cAMP/adenylyl cyclase pathway.

Hydrogen sulfide (H2S), traditionally known for its toxic effects, is now involved in regulating vascular tone. Here we investigated the vasoconstrictive effect of H2S on cerebral artery and the underlying mechanism. Sodium hydrosulfide (NaHS), a donor of H2S, concentration-dependently induced vasoconstriction on basilar artery, which was enhanced in the presence of isoprenaline, a ß-adrenoceptor agonist or forskolin, an adenylyl cyclase activator. Administration of NaHS attenuated the vasorelaxant effects of isoprenaline or forskolin. Meanwhile, the NaHS-induced vasoconstriction was diminished in the presence of 8B-cAMP, an analog of cAMP, but was not affected by Bay K-8644, a selective L-type Ca(2+) channel agonist. These results could be explained by the revised effects of NaHS on isoprenaline-induced cAMP elevation and forskolin-stimulated adenylyl cyclase activity. Additionally, NaHS-induced vasoconstriction was enhanced by removing the endothelium or in the presence of L-NAME, an inhibitor of nitric oxide synthase. L-NAME only partially attenuated the effect of NaHS which was given together with forskolin on the pre-contracted artery. In conclusion, H2S induces vasoconstriction of cerebral artery via, at least in part, cAMP/adenylyl cyclase pathway.

Vitamin K1 distribution following intravenous vitamin K1-fat emulsion administration in rats.

This study investigated vitamin K1 (VK1 ) distribution following intravenous vitamin K1-fat emulsion (VK1 -FE) administration and compared it with that after VK1 injection. Rats were intravenously injected with VK1-FE or VK1 . The organ and tissue VK1 concentrations were determined using high-performance liquid chromatography method at 0.5, 2 and 4 h to determine distribution, equilibrium and elimination phases, respectively. In the VK1-FE group, the plasma, heart and spleen VK1 concentrations decreased over time. However, other organs like liver, lung, kidney, muscle and testis, reached peak VK1 concentrations at 2 h. In the VK1 injection group, the liver VK1 concentrations were significantly higher than those in other organs at the three time points. However, VK1 concentrations in the other organs peaked at 2 h. In addition, in VK1-FE group, the heart, spleen and lung VK1 concentrations were significantly higher than those in the VK1 injection group at the three time points, and the liver VK1 concentration was significantly higher than that in the VK1 injection group at 4 h. The VK1 amount was greatest in the liver compared with the other organs. Thus, the liver is the primary organ for VK1 distribution. The distribution of VK1 is more rapid when injected as VK1-FE than as VK1 .

The severe adverse reaction to vitamin k1 injection is anaphylactoid reaction but not anaphylaxis.

The severe adverse reaction to vitamin K1 injection is always remarkable and is thought to result from anaphylaxis. Paradoxically, however, some patients administered vitamin K1 injection for the first time have adverse reactions. Using beagle dogs, the present study tested the hypothesis that the response to vitamin K1 is an anaphylactoid reaction. The results showed that serious anaphylaxis-like symptoms appeared in beagle dogs after the administration of vitamin K1 injection for the first time. The plasma histamine concentration increased, and blood pressure decreased sharply. After sensitization, dogs were challenged with vitamin K1 injection and displayed the same degree of symptoms as prior to sensitization. However, when the vitamin K1 injection-sensitized dogs were challenged with a vitamin K1-fat emulsion without solubilizers such asTween-80, the abnormal reactions did not occur. Furthermore, there was no significant change in the plasma immunoglobulin E concentration after vitamin K1 challenge. Following treatment with vitamin K1 injection, the release of histamine and ß-hexosaminidase by rat basophilic leukemia-2H3 cells as well as the rate of apoptosis increased. The Tween-80 group displayed results similar to those observed following vitamin K1 injection in vivo. However, the dogs in the vitamin K1-fat emulsion group did not display any abnormal behavior or significant change in plasma histamine. Additionally, degranulation and apoptosis did not occur in rat basophilic leukemia-2H3 cells. Our results indicate that the adverse reaction induced by vitamin K1 injection is an anaphylactoid reaction, not anaphylaxis. Vitamin K1 injection induces the release of inflammatory factors via a non-IgE-mediated immune pathway, for which the trigger may be the solubilizer.